Authors: Takashi Miura, Tika R. Malla, C. David Owen, Anthony Tumber, Lennart Brewitz, Michael A. McDonough, Eidarus Salah, Naohiro Terasaka, Takayuki Katoh, Petra Lukacik, Claire Strain-Damerell, Halina Mikolajek, Martin A. Walsh, Akane Kawamura, Christopher J. Schofield, Hiroaki Suga
Published: 2023-05-22
DOI: 10.1038/s41557-023-01205-1
Source: Full article
Abstractγ-Amino acids can play important roles in the biological activities of natural products; however, the ribosomal incorporation of γ-amino acids into peptides is challenging. Here we report how a selection campaign employing a non-canonical peptide library containing cyclic γ2,4-amino acids resulted in the discovery of very potent inhibitors of the SARS-CoV-2 main protease (Mpro). Two kinds of cyclic γ2,4-amino acids, cis-3-aminocyclobutane carboxylic acid (γ1) and (1R,3S)-3-aminocyclopentane carboxylic acid (γ2), were ribosomally introduced into a library of thioether-macrocyclic peptides. One resultant potent Mpro inhibitor (half-maximal inhibitory concentration = 50 nM), GM4, comprising 13 residues with γ1 at the fourth position, manifests a 5.2 nM dissociation constant. An Mpro:GM4 complex crystal structure reveals the intact inhibitor spans the substrate binding cleft. The γ1 interacts with the S1′ catalytic subsite and contributes to a 12-fold increase in proteolytic stability compared to its alanine-substituted variant. Knowledge of interactions between GM4 and Mpro enabled production of a variant with a 5-fold increase in potency.